RSV appears in predictable yearly outbreaks. Annual outbreaks of lower respiratory tract disease in young children have been noted since at least the early 1940's (1). RSV was implicated as the major cause of these outbreaks soon after its discovery in 1956 (2, 3). RSV infects adults as well as infants, and causes serious lower respiratory tract disease primarily in very young infants, children with pulmonary or cardiac disease, the immunologically compromised and the elderly (4). RSV infection is responsible for 40% to 50% of cases of children hospitalized with bronchiolitis and 25% of children with pneumonia (5). The number of cases requiring hospitalization in 1993 has been estimated at 91,000 with a cost of approximately $300,000,000 (5). Spread of the virus in hospitals is a particularly serious problem. When RSV infections are present in a hospital, 20% to 45% of infants may acquire a nosocomial RSV infection (6). Premature infants and those hospitalized for cardiac or pulmonary diseases are thus placed at acute risk of developing lower respiratory tract disease. In a study of children with congenital heart disease, 21% of RSV infections were acquired nosocomially (6).
To date, an effective vaccine against RSV has not been developed. In lieu of an active vaccine to protect high risk patients, especially infants, passive application of antibody may serve to protect these children during periods of known exposure. Intravenous treatment with immunoglobulin (IgG) containing anti-RSV activity is being tested in clinical trials (7, 8). While intravenous IgG might prevent lower respiratory tract disease, the evidence suggests that large doses and volumes of this material are required. Such treatment is not without potential adverse effects, including volume overload and circulatory failure.
In humans, upper airway infection generally precedes involvement of the lower respiratory tract (4). A study of modes of transmission shows that the virus is spread via fomites and self-inoculation of the nose or eyes rather than by aerosol, suggesting that the infection does not initiate in the lower respiratory tract (14). Viral infection is normally limited to the respiratory tract epithelium, and cell-to-cell spread is probably via secretions and cell-cell fusion (4). Fused cells can be recovered from lung aspirates of infected patients (4), but the importance of syncytium formation in pathogenesis or viral spread is not known.
None of the current approaches to prophylaxis of RSV focuses on the prevention of initial stages of infection in the upper respiratory tract. Natural immunity in this compartment of the respiratory tract is mediated by IgA antibodies in the nasal secretions.
The immune response to RSV infection is short-lived. This allows repeated infection to occur in adults and children. In an adult challenge study, 40% of the subject could be infected 3 times with the same challenge strain over a period of 26 months (15). Up to 75% of children infected during their first season of RSV exposure are reinfected in their second season (16), although severe disease was uncommon after the initial infection. Circulating anti-RSV antibody can be protective when present in sufficient quantity, but its importance has been difficult to resolve. In animals, human IgG or specific monoclonal antibodies administered parenterally can protect against replication of the virus in the lung (9, 17-19). High levels of circulating anti-RSV antibody protects primarily the lower respiratory tract (9). Moreover, the role of secretory antibody in protection against RSV has not been clearly established, but it appears that it may be an important mediator of the upper airway immunity. The titer of neutralizing antibody in nasal secretions correlates with decreased virus shedding and protection against disease in adult volunteers challenged with RSV (20, 21). A decrease in viral shedding also correlates with the appearance of anti-RSV secretory IgA (sIgA) in nasal secretions of infants (22). However, not all of the neutralizing activity of nasal secretions is due to antibody (22). A correlation between nasal anti-RSV antibody level and protection against infection or severe disease has not been demonstrated in human infants (15, 22-25), but in animals, mucosal immunization protects against nasal infection (26-27).
RSV is an enveloped, negative strand RNA virus belonging to the genus Pneumovirus of the Paramyxoviridae family (10, 11). Two glycoproteins, 90 kD and 68 kD, are exposed on the surface of the virion. The 90 kD heavily glycosylated G protein is responsible for binding of virus particles to target cells (12). The 68 kD F protein mediates fusion of the viral envelope with the cell membrane and syncytium formation (13). The F and G surface glycoproteins referred to above appear to be the primary protective antigens, with the nucleoprotein N and the envelope protein M2 having minor protective activity. Neutralizing and fusion-inhibiting monoclonal antibodies have been mapped to specific domains of F glycoprotein (9, 28-31). Monoclonal antibodies against the G glycoprotein are less likely to neutralize virus than those against the F glycoprotein and do not have fusion inhibiting activity (32-34). The amino acid sequence of F glycoprotein is approximately 90% conserved between the RSV subgroups responsible for human infection (35). Conserved epitopes include some that mediate neutralization and fusion inhibition ( 19, 35). The G glycoprotein which is primarily responsible for differences between subgroups A and B is only 53% conserved between the two subgroups (36). Immunization with the vaccinia virus recombinant expressing N or M2 induces a minor protective response in mice (37, 38). This response may be due primarily to CTL activity, since anti-N monoclonal antibody does not protect when passively administered to mice (39). Moreover, N has been shown to be a CTL target in mice and humans (37).
An early vaccine consisting of formalin-inactivated alum-adsorbed RSV elicited neutralizing and complement-fixing serum antibody in a clinical trial. However, vaccinated children were not protected and had more severe lower respiratory tract disease upon subsequent natural infection (40). The reason for the enhanced disease has not been fully explained. Cotton rats immunized with formalin-inactivated RSV developed a similar pathological response (41), providing a method of testing the safety of new vaccines.
Efforts have focused in the past on developing attenuated live virus vaccines. To date, those vaccines have been found to be ineffective (42), insufficiently attenuated (43, 44), or genetically unstable (45, 46). More recent efforts have focused on the RSV surface glycoproteins F and G. Immunization with purified F glycoprotein has been shown to be effective in cotton rats and is currently in clinical trials (47-48). However, some preparations of F glycoprotein have been shown to cause enhanced lung pathology upon subsequent RSV infection in cotton rats (49). Recombinant chimeric FG glycoprotein produced in a baculovirus expression system elicits a protective immune response in cotton rats when given parenterally (50). As with F glycoprotein alone, FG vaccine was also shown to cause some enhanced pulmonary pathology in cotton rats (51, 52). Vaccinia virus recombinants expressing F, G or M2 envelope protein, or the nucleoprotein N, have been tested in several animal models. F and G recombinants have shown the most promise, inducing protective immunity in mice (53, 54), cotton rats (55) and owl monkeys (56). The response in chimpanzees however was markedly lower (57). Adenovirus is also being examined as a vector for RSV F glycoprotein (58).
A need exists, therefore, for effective approaches to the prevention of RSV disease. The present invention seeks to fill that need.